GC numbers and size are equally increased in AID−/− and AID−/−µS−/− animals and AID−/−µS−/− and μS−/− mice have atypical IgMhiIRF4+CD138− plasmablast-like cells. Spleen sections of age-matched WT C57Bl/6, AID−/− (AID), μS−/−, and AID−/−µS−/− mice were analyzed by confocal microscopy. (A) Ki67 (blue) highlight clusters of proliferating GC B cells adjacent to CD21+ follicular dendritic cells (green) in the follicles constituted by CD23+ B cells (red). (B) The proportion of follicles seeded with GC was determined for each of the 4 spleens taken from 4 mice in each group. For each spleen section taken from each of the 4 spleens of the same group, the numbers of follicles and GCs were counted on 5 nonoverlapping images similar to that in A, representing in total 1.6 mm2/spleen. Numbers of GL7+FAS+ GC B cells determined by FACS analysis of spleens (C) and mLNs (D) from the 4 groups of mice. (E) In the top panels, IRF-4 (red) identifies plasmablasts and PCs. Unswitched PCs co-stained for IgM (blue) appear in pink, whereas switched IRF-4+IgM-IgD− PCs are red. In WT and AID−/− spleens, PC clusters are in the red pulp, whereas many IgMhiIRF-4+ PC-like cells (pink) are scattered throughout the follicles (IgD+ area in green in WT, AID−/−, and μS−/−, or cyan in AID−/−µS−/−), MZ, and red pulp. On the bottom panels, Ki67 (green) highlights clusters of proliferating cells in the red pulp of AID−/−µS−/− and μS−/− mice. Switched IgM-CD138+ PC in red, visible only in WT spleens, and unswitched PCs positive for IgM (blue) and CD138 (red), in pink, seen in both WT and AID−/− spleens are absent from AID−/−µS−/− and μS−/−. (F) Expression of IRF-4, Blimp-1, and XBP-1 mRNA was quantified in spleen sections from the 4 groups of mice by real-time reverse transcription-PCR, relative to the level of β2-microglobulin mRNA used as reference gene. Representative confocal images are shown; results are representative of 3 independent experiments; *P ≤ .05; **P ≤ .01; ***P ≤ .005. White bars on confocal images represent 200 μm.
